Dataset Information

The Cancer Genome Atlas (TCGA) Consortium Integrated DNA Methylation Analysis

ABSTRACT: Aberrant hypermethylation of CpG dinucleotides located in CpG islands within the promoters of key cancer genes is an epigenetic abnormality associated with heritable transcriptional gene silencing and inactivation in cancer. The genes involved include important tumor suppressors affecting key pathways for tumor initiation and progression. These methylated sequences can serve as potentially valuable markers for cancer risk assessment, diagnosis, prognosis, and prediction of therapeutic responses. In addition, many key cancer genes may be targeted by both epigenetic and genetic alterations and, thus epigenetic analysis can help focus the search for mutations, and vice versa. Studies of major cancer types suggest that any individual patient’s tumor may harbor at least 300 or more DNA hypermethylated genes. In TCGA, a pilot project is underway to begin defining these genes for GBM via genomic approaches. The approach in the epigenetic pilot is a two-tiered one which, first, involves pharmacological treatment of both well established human GBM cell lines, and a cell line grown as a neurosphere to enrich for tumor propagating cells, with a DNA methylation inhibitor (5-aza-2’-deoxycytidine, DAC) or a histone deacetylation inhibitor (trichostatin A) followed by an expression transcriptome analysis as previously described (Schuebel et. al.). This has resulted in identification of more than 3,700 total candidate genes. In the second tier, the top candidates are then analyzed on a custom Illumina GoldenGate array with the capacity to monitor methylation at a single CpG dinucleotide in the CpG islands of 1,498 gene promoters for the high throughput analysis of TCGA GBM samples. Keywords: Microarray, Hypermethylome, DNA-hypermethylation, DAC, TSA, Epigenetic, TCGA, The Cancer Genome Atlas, GBM, Glioblastoma, Glioblastoma multiforme, Brain

ORGANISM(S): Homo sapiens

PROVIDER: GSE11233 | GEO | 2009/01/01

SECONDARY ACCESSION(S): PRJNA106731

REPOSITORIES: GEO

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Project description:Estrogen signaling and epigenetic modifications, in particular DNA methylation, are involved in regulation of gene expression in breast cancers. Here we investigated a potential regulatory cross-talk between these two pathways by identifying their common target genes and exploring potential underlying molecular mechanisms in human MCF7 breast cancer cells. Principal Findings: Gene expression profiling revealed that the expression of approximately 150 genes was influenced by both 17β-estradiol (E2) and a hypomethylating agent 5-aza-2’-deoxycytidine (DAC). Based on gene ontology (GO), CpG island prediction analysis and previously reported estrogen receptor (ER) binding regions, we selected six genes for further analysis (BTG3, FHL2, PMAIP1, BTG2, CDKN1A and TGFB2). GO analysis suggests that these genes are involved in intracellular signaling cascades, regulation of cell proliferation and apoptosis, while CpG island prediction of promoter regions reveals that the promoters of these genes contain at least one CpG island. Using chromatin immunoprecipitation, we show that ERα is recruited to CpG islands in promoters, but neither in an E2- nor in a DAC-dependent fashion. DAC treatment reactivates the expression of all selected genes although only the promoters of BTG3 and FHL2 genes are methylated, with E2 treatment showing no effect on the methylation status of these promoters. Conclusions: We identified a set of genes regulated by both estrogen signaling and DNA methylation. However, our data does not support a direct molecular interplay of mediators of estrogen and epigenetic signaling at promoters of regulated genes. The aim of the study was to identify genes regulated by estrogen signaling and DNA methylation, using microarray approach. We compared the effects of E2 and DAC on global gene expression profiles in MCF7 cells. By comparing C_E2 (MP5-MP8 average) with C_EtOh samples (MP1-MP4 average), 802 genes were identified as up-regulated by E2, while 851 genes were identified as down-regulated by E2. 1017 genes were identified as up-regulated by DAC, by comparing A_EtOh (MP9-MP12 average) samples with C_EtOh, suggesting that DNA methylation is involved in their regulation. To identify possible common targets, we have compared the DAC up-regulated genes with E2-regulated genes. 88 annotated genes are found to be up-regulated by both E2 and DAC, suggesting that E2 has a hypomethylation-like effect on the regulation of these genes. 58 annotated genes are found to be down-regulated by E2 and up-regulated by DAC, suggesting that E2 has a hypermethylation-like effect on the regulation of these genes.

Project description:The model describing that aberrant CpG island (CGI) methylation leads to transcription repression of tumor suppressor genes and thereby is implicated in tumor progression has been established in many cancers. However, recent studies indicated aberrantly hypermethylated genes in multiple cancers are already repressed in pre-cancerous tissues despite their promoters are hypomethylated. Here, we hypothesized that the occurrence of CGI promoter hypermethylation in cancers are associated with Polycomb-repressive complex and the associated H3K27me3 mark in pre-cancerous tissues. By using a ChIP-BS-seq technology that examines methylation of the DNA fragments precipitated by the antibodies to histone modifications, we provided direct evidences showing that genes highly enriched with H3K27me3 marks both in cancer and normal cells became aberrantly hypermethylated in CGI promoters in cancer cells in comparison with normal cells. Furthermore, we confirmed that these genes consistently were significantly hypermethylated in TCGA primary cancer in comparison with normal tissues. Thus, we provided direct evidences supporting that the presence of H3K27m3 may serve as a guide to promoter hypermethylation. This will spur future work on epigenetic signature of combined histone and DNA methylation that could define a cancerM-bM-^@M-^Ys epigenetic abnormalities, therefore helping distinguish subtypes of cancers and aiding future diagnosis and therapeutics of cancers. We applied ChIP-BS technology to examine H3K27me3 marks, which are catalyzed by the SET domain histone methyltransferase EZH2 and have a repressive function with 50bp pair-end sequencing. found H3K27me3 marks were enriched preferentially at CpG islands, (+/-500) transcription start sites (TSSs) and exons in two GC cell lines (BGC-823 and AGS). In YH cells, H3K27me3 marks were only preferentially enriched at CpG islands. In contrast, Hela cells presented a reverse pattern with highest H3K27me3 enrichment in intergenic regions. To confirm this result in Hela cells, we performed two independent replicates of ChIP-Seq and ChIP-BS-seq. Cause of useful was relative small. we still sequenced one 100bp pe reads replicate for H3K4me3 and two replicate for H3K27me3 ChIP-BS-seq.

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The Cancer Genome Atlas (TCGA) Consortium Integrated DNA Methylation Analysis

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